半滑舌鳎脑芳香化酶基因cDNA克隆及表达分析

为研究脑型芳香化酶(P450aromB)在半滑舌鳎性别分化中的作用,采用同源克隆策略,从半滑舌鳎脑分离了2184bp长的脑型芳香化酶的全长cDNA,该基因编码498个氨基酸。氨基酸序列和系统发育分析表明,P450aromB属于脑型P450arom,P450aromB的氨基酸序列与其他鱼类脑型P450arom的同源性较高(48.3%-66.1%),与性腺型P450arom的同源性较低(34.2%-49.9%),与自身的性腺型芳香化酶同源性为45.1%。RT-PCR分析表明:P450aromB mRNA的表达具有明显组织特异性,P450aromB只在性腺、脑、鳃和皮肤中表达,且脑中表达量远高于性腺,而在雌雄鱼的其他组织中都不表达。经过甲基睾酮浸浴处理和高温诱导半滑舌鳎由雌性性反转为雄性后,脑中P450aromB的表达量降低,这些结果表明P450aromB参与了半滑舌鳎的性腺分化和性别决定过程。

Molecular Cloning, Characterization and Expression Analysis of Brain P450arom in Half-smooth Tongue-sole, Cynoglossus semilaevis Gunther

s:The 2184 bp full-length brain P450 aromatase (termed P450aromB) cDNA encoding a protein of 498 amino acid residues was isolated from the brain of the half-smooth tongue-sole (Cynoglossus semilaevis) by using homologous cloning to clarify the role in sex differentiation. Sequence and phylogenetic analyses showed that the tongue sole P450aromB belonged to brain P450arom subfamilies. The putative P450aromB amino acid residues shared 48.3%- 66.1% sequence affinity with other fish brain P450arom, and shared 34.2%-49.9% sequence affinity with ovarian P450arom of other fish. The RT-PCR analysis revealed that P450aromB mRNA was expressed at a high proportion in the brain and gills but less in the gonads and skin, but not in other tissues. However, the P450aromB decreased in the brain of sex-reversed males, treatment by Methyltestosterone or at high temperatures. These results suggested that the P450aromB is involved in the course of gonad development and sex determination in tongue soles.